Advances in human reproductive medicine require in-depth knowledge about the structure and function of gamete proteins. In our recent proteomic analysis of the accessory structures of mouse sperm tails, we demonstrated that important enzymes of adenine nucleotide metabolism are present in the flagellum. Besides basal sperm motility, key sperm functions such as capacitation and hyperactivated motility rely on the use of adenine nucleotides as energy sources and as signaling molecules. However, the mechanisms integrating adenine nucleotides into various pathways of synthesis, degradation, and signaling in sperm are poorly understood. Thus, the broad, long-term goal of this project is to determine how the relative roles of ATP, ADP, and AMP influence sperm motility. We propose that the adenylate energy charge in a given region of the sperm flagellum impacts the quality and vigor of movement in each microenvironment through the energy provided by ATP and modulated by signaling mediated via ATP, ADP, and AMP. To address these issues, we propose three specific aims. Aim 1 is to determine how sperm motility is regulated by adenylate kinase through the interconversion of ADP to ATP plus AMP. The hypothesis to be tested is that the adenylate energy charge within a given sperm cell affects the quality of motility and underlies the changes associated with hyperactivation. Aim 2 is to Identify AK proteins of sperm and the flagellar proteins that interact with them. The hypothesisto be tested is that sperm AKs form regulatory complexes with other proteins of the flagellum. Aim 3 is to determine the regulatory pathway(s) utilized by AMP to modify sperm motility. The hypothesis to be tested is that AMP-sensitive enzymes are important regulators of sperm physiology. We propose that adenine nucleotides serve multiple roles as primary energy stores (ATP), supplemental energy (ADP), and energy gauge (AMP) to regulate sperm metabolism and motility. PUBLIC HEALTH RELEVANCE: Sperm must have proper motility to fertilize an egg. Understanding the roles of adenine nucleotides in regulating the rate, amplitude, and waveforms of the sperm flagellum may provide new ways to assist couples experiencing infertility and to develop new forms of contraception. Information gained from these studies may aid in the understanding of globally important diseases/infections involving flagella (e.g., polycystic kidney disease, trypanosomes).